| 高肺转移人源骨肉瘤细胞系的建立及鉴定 |
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| 引用本文： | 支文兰,常君丽,胡少朴,王晓波,赵福来,孙星媛,马小平,杨燕萍.高肺转移人源骨肉瘤细胞系的建立及鉴定[J].中国癌症防治杂志,2021,13(4):394-399. |
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| 作者姓名： | 支文兰 常君丽 胡少朴 王晓波 赵福来 孙星媛 马小平 杨燕萍 |
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| 作者单位： | 上海中医药大学附属龙华医院 上海中医药大学脊柱病研究所 教育部筋骨理论与治法重点实验室 |
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| 基金项目： | 国家重点研发计划项目（2018YFC1704300；2020YFE021600）；国家自然科学基金项目（81973877；81674006） |
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| 摘 要： | 目的 建立具有高肺转移能力的人源骨肉瘤细胞系，为探索骨肉瘤肺转移的发生机制提供研究模型。方法 将GFP和荧光素酶标记的人源骨肉瘤143B细胞重悬到MEM培养基并与Matrigel胶按1∶1比例混匀，制成浓度为1×107/mL的细胞悬液。取10 μL注射到4周龄BALB/c 裸鼠的左侧胫骨内，建立胫骨内原位注射肺转移瘤模型（n=6）。收集肺转移结节制成单细胞悬液，在完全培养基中贴壁培养，并用新霉素（G418）抗性筛选后扩增，再次接种于裸鼠胫骨内；采用同样方法在体内循环2次后获得的细胞命名为143B-HLM。采用体外实时细胞迁移实验检测细胞迁移情况，RT-qPCR和Western blot检测迁移浸润关键分子标志物的表达情况，同时观察肺转移瘤模型肺部肿瘤结节的形成情况。结果 与亲本人源骨肉瘤143B细胞相比，建立的143B-HLM细胞体外迁移能力增强，波形蛋白（Vimentin）和锌指转录因子（Slug）的蛋白表达量增加（均P<0.05），Vimentin、Slug、锌指转录抑制因子（Snail）和凋亡抑制基因（Survivin）的mRNA表达水平也升高（均P<0.05）。与注射143B细胞的裸鼠相比，裸鼠胫骨内原位注射143B-HLM细胞的荷瘤模型中，原位肿瘤体积减小（P<0.05），肺部肿瘤转移荧光信号显著增强，肺转移的裸鼠数目也更多（P<0.01）。结论 骨肉瘤肺转移细胞经体内循环筛选及分离，成功建立了稳定的具有高肺转移能力的骨肉瘤细胞系143B-HLM。
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Establishment and identification of human osteosarcoma cell line with high lung metastasis |
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| ZHI Wenlan,CHANG Junli,HU Shaopu,WANG Xiaobo,ZHAO Fulai,SUN Xingyuan,MA Xiaoping,YANG Yanping.Establishment and identification of human osteosarcoma cell line with high lung metastasis[J].Chinese Journal of Oncology Prevention and Treatment,2021,13(4):394-399. |
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| Authors: | ZHI Wenlan CHANG Junli HU Shaopu WANG Xiaobo ZHAO Fulai SUN Xingyuan MA Xiaoping YANG Yanping |
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| Abstract: | Objective To establish a human osteosarcoma cell line with high lung metastasis ability and provide a research model for explore the mechanism of lung metastasis of osteosarcoma. Methods The human osteosarcoma 143B cells labeled with GFP and luciferase were resuspended into a MEM medium and mixed with Matrigel gel (1∶1) to make a cell suspension (1×107/mL) , and 10 μL of the suspension was injected into the left tibia of 4-week-old BALB/c nude mice to establish a model of lung metastasis by in situ injection into the tibia (n=6) . The lung metastasis nodules were collected to make a single-cell suspension, cultured adherently in a complete medium, and amplified with neomycin (G418) resistance screening, then inoculated into the tibia of nude mice again. The cells obtained after circulating twice in vivo by the same method were named 143B-HLM. The cell migration was detected by the vitro real-time cell migration assay, the expression of key molecular markers of cell migration and infiltration was detected by RT-qPCR and Western blot, and lung tumor nodule formation in lung metastatic tumor model was observed. Results Compared with parental human osteosarcoma 143B cells, 143B-HLM cells had higher migratory capacity in vitro; the protein expression of Vimentin and Slug, and the mRNA expression levels of Vimentin, Slug, Snail, and Survivin were increased (all P<0.05) . Compared with nude mice injected with 143B cells, the tumor-bearing model in which 143B-HLM cells were injected in situ into the tibia of nude mice, the tumor volume in situ was decreased (P<0.05), the fluorescence intensity of lung tumor metastasis was significantly increased, and the number of lung metastasis was also increased (P<0.01). Conclusions A stable osteosarcoma cell line 143B-HLM with high lung metastatic capacity was successfully established through in vivo circulation screening and isolation of osteosarcoma metastatic cells. |
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| Keywords: | Osteosarcoma cell line 143B-HLM High lung metastasis cell line |
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